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>Stereospecificity of hydrogen transfer by pyridine nucleotide-dependent enoyl reductases in fatty acid synthesis: Studies with enzymes obtained from developing castor bean seeds andChlorella vulgaris
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Stereospecificity of hydrogen transfer by pyridine nucleotide-dependent enoyl reductases in fatty acid synthesis: Studies with enzymes obtained from developing castor bean seeds andChlorella vulgaris
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机译:Stereospecificity of hydrogen transfer by pyridine nucleotide-dependent enoyl reductases in fatty acid synthesis: Studies with enzymes obtained from developing castor bean seeds andChlorella vulgaris
The mechanism of hydrogen incorporation into fatty acids was investigated with fatty acid synthetase systems from developing castor bean seeds andChlorella vulgaris. Fatty acids synthesized in the presence of D2O or stereospecifically deuterium-labeled NADPH or NADH were isolated and analyzed by mass spectrometry to examine the localization of deuterium atoms in the molecule. The stereospecificities ofβ-ketoacyl-acyl carrier protein (ACP) reductase and enoyl-ACP reductase for reduced pyridine nucleotide were determined with acetoacetyl-ACP and crotonyl-ACP as substrates. The products were also analyzed by gas chromatography-mass spectrometry. The following results were obtained:β-Ketoacyl-ACP reductases from both castor bean seeds andC. vulgarisused the B-side hydrogen of NADPH.Enoyl-ACP reductase fromC. vulgarisrequired NADH for the activity.Enoyl-ACP reductase from castor bean seeds used the A-side hydrogen of NADPH, whereas that fromC. vulgarisused the B-side hydrogen of NADH.When stearate was synthesized with the crude fatty acid synthetase fraction from castor bean seeds, hydrogen atoms from water were found on the even-numbered methylene carbon atoms (two hydrogen atoms per carbon atom) and some were found on the odd-numbered methylene carbon atoms. Hydrogen atoms from the B-side of NADPH were found on the odd-numbered methylene carbon atoms (one hydrogen atom per carbon atom). Hydrogen atoms from the A-side of NADPH were also found on the odd-numbered methylene carbon atoms, but the number of incorporated hydrogen atoms was less than expecte
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